This sequence change replaces leucine, which is neutral and non-polar, with phenylalanine, which is neutral and non-polar, at codon 333 of the CFTR protein (p.Leu333Phe). This variant is present in population databases (rs193922533, gnomAD 0.002%). This missense change has been observed in individual(s) with clinical features of CFTR-related conditions (PMID: 28502372, 33374015). ClinVar contains an entry for this variant (Variation ID: 35895). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt CFTR protein function with a positive predictive value of 80%. In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance.

The p.L333F variant (also known as c.997C>T), located in coding exon 8 of the CFTR gene, results from a C to T substitution at nucleotide position 997. The leucine at codon 333 is replaced by phenylalanine, an amino acid with highly similar properties. This alteration has been detected once in a cohort of individuals with acute recurrent and chronic pancreatitis (Giefer MJ et al. J. Pediatr., 2017 07;186:95-100). The p.Leu333Phe variant was identified in conjunction with p.Phe508del in one of 71 male patients with infertility, although phase was not confirmed (Rudnik-Schöneborn S et al. Hum Reprod, 2021 Feb;36:551-559). This amino acid position is highly conserved in available vertebrate species. In addition, the in silico prediction for this alteration is inconclusive. Based on the available evidence, the clinical significance of this variant remains unclear.

Variant summary: CFTR c.997C>T (p.Leu333Phe) results in a non-conservative amino acid change located in the ABC transporter type 1, transmembrane domain (IPR011527) of the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 8e-06 in 251260 control chromosomes. The available data on variant occurrences in the general population are insufficient to allow any conclusion about variant significance. c.997C>T has been reported in the literature in compound heterozygosity with the common pathogenic variant p.Phe508del in at least one individual affected with azoospermia or oligospermia (exact phenotypic details not provided, e.g. Rudnik-Schoneborn_2021). In addition the variant was reported in an individual with pancreatitis without strong evidence for causality (e.g. Giefer_2017). These reports do not provide unequivocal conclusions about association of the variant with Cystic Fibrosis. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. No clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014. Based on the evidence outlined above, until additional information becomes available, the variant was classified as uncertain significance.

This variant was identified in 1 patient with a clinically confirmed diagnosis of cystic fibrosis. The variant was classified in the context of a project re-classifying variants in the German Cystic Fibrosis Registry (Muko.e.V.). Link: https://www.muko.info/angebote/qualitaetsmanagement/register/cf-einrichtungen/mukoweb. Criteria applied: PM3, PM2_SUP, PP3